1. Field of the Invention
The present invention relates generally to biomedical compositions and methods for treating diseases, disorders and conditions affecting connective tissues. In particular, the present invention provides compositions and methods for enhancing elastic fiber content by promoting molecular mechanisms of elastogenesis.
2. Description of the Related Art
The extracellular matrix (ECM) plays an important role in the development and maintenance of vertebrate tissues, including the organization of various ECM components to provide structurally and functionally significant connective tissue compartments (reviewed in e.g., Hay, 1981 J. Cell Biol. 91:205s; Dallas et al., 2006 Curr. Top. Dev. Biol. 75:1; Ushiki, 2002 Arch. Histol. Cytol. 65:109). An important but incompletely characterized component of the ECM comprises the elastic fibers, a system of microfibrils that contains the protein elastin and other polypeptides in a framework that confers elasticity and resilience to selected tissues, including blood vessels, cardiac tissues including heart valves, lung, gastrointestinal and urogenital tracts, cartilage, ligaments, tendons and skin (Kielty et al., 2002 J. Cell Sci. 115: 2817; Cleary and Gibson, 1996 in Extracellular Matrix (Comper, W. D. ed.) Vol. 2, pp. 95-140, Harwood Academic Publishers, Amsterdam; Mecham and Heuser, 1991 in Cell Biology of the Extracellular Matrix (Hay, E. D., ed.) 2nd Ed., pp. 79-109, Plenum Publishing Corp., New York; Pasquali-Ronchetti et al., 1997 Microsc. Res. Tech. 38:428).
While many ECM components are synthesized throughout the lifetimes of most vertebrates, elastic fiber biosynthesis, or elastogenesis, takes place largely in developing tissues (Davis, 1993 Histochem. 100:17) with little or no de novo elastic fiber synthesis being seen in mature tissues. As such, the elastic fiber compartment of the ECM is not replenished following elastic fiber degradation due to aging or disease, or subsequent to tissue damage incurred in the course of injury, surgery or other clinical insults. The resultant loss of tissue elasticity and resiliency may exacerbate the disease, for example, by impairing the physiological function of the affected tissue and/or predisposing the afflicted individual to further tissue damage. The benefits of a functional elastic fiber system in an individual may thus be compromised by disease and/or related sequelae, for instance, by aneurysms, restenosis lesions, emphysema, or Marian syndrome, by skin wounds or aging-associated damage such as skin wrinkling, and/or by genetic defects linked to the elastin gene (see, e.g., Robb et al., 1999 Mol. Biol. Cell 10:3595 and references cited therein).
As known in the art, certain naturally occurring cell surface glycoconjugates act as elastogenic inhibitors by binding to the cell surface elastin binding protein (EBP) and disrupting EBP elastogenic activities. For example, several recent studies have highlighted the reciprocal relationship between elastogenesis and matrix proteoglycan content of tissues (e.g., Wight and Merrilees, 2004 Circ. Res. 94:1158; Hinek et al., 1991 J. Clin. Invest. 88:2083). Elastic fibers are generally absent or depleted in matrices rich in chondroitin sulphate (CS)-containing proteoglycans, and correspondingly increased in matrices depleted of CS proteoglycans (Kolodgie et al., 2002 Arterioscler. Thromb. Vasc. Biol. 22:1642-1648). Several lines of evidence point to the premature detachment of the elastin binding protein (EBP) from the cell surface as the mechanism by which the assembly of elastic fibers is disrupted (Hinek et al., 2000 Am. J. Hum. Genet. 66:859-872). There is compelling evidence that this detachment is due to the high concentration CS-containing proteoglycans, notably versican, found in pericellular coats of matrix rich tissues (Huang et al., 2006 Circ. Res. 98:370). EBP is an inactive splice variant of β galactosidase that binds both tropoleastin monomers and galactosugars such as CS (e.g., Hinek et al., 1993 J. Clin. Invest. 91:1198). Premature release of EBP from the cell surface occurs when the binding pocket for galactosugars is occupied (Hinek et al., 1991 J. Clin. Invest. 88:2083).
Proteoglycan rich matrices usually contain two major proteoglycans, versican and biglycan. (For versican see, e.g., Wight and Merrilees, 2004 Circ. Res. 94:1158; Wight, 2002 Curr. Opin. Cell Biol. 14:617; Yao et al., 1994 Matrix Biol. 14:213; Wight, The vascular extracellular matrix. In: Fuster V., Topol E., Nabel E., eds., Atherosclerosis and Coronary Artery Disease, 2004 Lippincott Williams and Wilkins, Philadelphia, Pa.; Nikkari et al., 1994 Am. J. Pathol. 144:1348; for biglycan see, e.g., Schwarz et al., 1990 J. Biol. Chem. 265:22023; Itabashi et al., 2005 Connect Tissue Res. 46:67; Tufvesson et al., 2002 Eur. J. Biochem. 269:3688; Grande-Allen et al., 2004 Glycobiol. 14:621; Theocharis et al., 2002 Atherosclerosis 165:221; Stanescu 1990 Sem. Arth. Rheum. 20(3 Suppl. 1):51). Versican, with its multiple CS GAG chains, has been shown to be an effective inhibitor of elastogenesis (Huang et al., 2006 Circ. Res. 98:370). It is unclear, however, whether biglycan, a small leucine-rich proteoglycan (SLRP) which possesses two GAG chains containing chondroitin and dermatan sulphates, may also play a role in modulating elastic fiber formation.
For example, biglycan core protein was shown to be capable of binding to tropelastin and to elastic fiber microfibrils (Reinboth et al., 2002 J. Biol. Chem. 277:3950), and in a kidney injury model, biglycan stimulated expression of fibrillin-1, a major component of the microfibrils that form the scaffold on which tropoelastin is deposited (Schaefer et al., 2004 Am. J. Pathol. 165:383). Biglycan gene expression also decreased in abdominal aortic aneurysms where elastin was disrupted and fragmented (Theocharis et al., 2002 Atherosclerosis 165:221). On the other hand, biglycan stimulated cellular proliferation and migration, and induced cell elongation, features that are associated with a non-elastogenic phenotype (Shimizu-Hirota et al., 2004 Circ. Res. 94:1067; Kinsella et al., 2004 Crit. Rev. Euk. Gene Expr. 14:203).
Existing strategies to encourage ECM remodeling typically involve administering compositions containing pre-formed ECM components or materials derived therefrom, but generally lack any demonstration of induced ECM biosynthesis or of de novo elastic fiber formation.
For example, JP2004/250395 describes a topically applied composition containing collagen, elastin, chondroitin sulfate and hyaluronic acid to counteract aging in skin, bone and hair. WO2004/012665 describes an orally administered composition containing chondroitin sulfate, glucosamine, methylsulfonylmethane (MSM) and for disorders of joints and connective tissue. CA2518794 describes a lipid for topical application to deep airway alveoli to reduce lung surface tension in emphysema. CA2520057 describes plant flavonolignans (“silymarins”) purported to promote type I collagen and/or elastin production in skin. WO2005/082386 describes cationic iron and magnesium salts used with digested elastic tissue to stimulate smooth muscle cell (SMC) elastogenesis and inhibit SMC proliferation, and to promote skin ECM deposition and dermal fibroblast proliferation. US2004/0146539 describes topical formulations containing vitamins, metabolites and/or plant extracts for cosmetic improvement to skin, including reducing wrinkles and other effects of aging. FR2847816 describes topical formulations containing an artificial peptide and vitamins (other than vitamin C) or nutrients for cosmetic improvement to skin, including reducing wrinkles and other effects of aging.
WO 0191700 describes small peptides derived from the amino acid sequence of elastin, and related structures, for administration orally, topically or via other parenteral routes to enhance connective tissue elasticity; peptide-coated vascular stents are also contemplated for improving flexibility and elasticity of blood vessels. WO99/45942 teaches the use of metastatin, a hyaluronan-binding complex derived from proteolytically digested cartilage, to inhibit cancer and angiogenesis, but is silent with respect to elastogenesis.
Transgenic approaches to promote elastic fiber biosynthesis in vitro have been described including transfection of cells with recombinant expression constructs encoding the elastin precursor subunit polypeptide tropoelastin (Robb et al., 1999 Mol. Biol. Cell 10:3595), and the V3 splice variant of the versican proteoglycan polypeptide (US 2004/0213762), which lacks chondroitin sulfate proteoglycan (CSPG) carbohydrate moieties but contains an intact hyaluronan-binding domain (Wight and Merrilees, 2004 Circ. Res. 94:1158). In this regard it is not completely understood what is the mechanism by which V3 may promote elastogenesis, in particular where both V3 and the CSPG-containing versican isoforms may be capable of influencing elastin fiber assembly at the cell surface by virtue of their binding interactions with cell surface hyaluronic acid (Wight and Merrilees, 2004 Circ. Res. 94:1158; Hinek et al., 2004 Am. J. Pathol. 164:119; Hinek et al., 2000 Am. J. Hum. Genet. 66:859; Hinek, 1996 Biol. Chem. 377:471; Hinek, 1994 Cell Adhes. Commun. 2:185).
Clearly there is a need for improved compositions and methods that exploit heretofore unrecognized molecular interactions to promote elastogenesis. The presently disclosed invention embodiments fulfill such a need and offer other related advantages.